Purification of recombinant proteins with an example of tumor necrosis factor thymosin-a1

Hybrid protein, cancer necrosis factor thymosin-a1 (TNF-T), when synthesizing in strain-producer of Escherichia coli SG200-50 with plasmid pThy315, was a part of 'inclusion bodies' mostly in the form of a high-molecular complex with other proteins due to the S-S bonds formation. An approac...

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Bibliographic Details
Published in:Applied biochemistry and microbiology 2010-03, Vol.46 (2), p.226-229
Main Authors: Fedorov, T V, Korobov, VI, Nazarov, V G, Smolkina, A E, Shmelev, V A
Format: Article
Language:English
Subjects:
Escherichia coli
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Summary:Hybrid protein, cancer necrosis factor thymosin-a1 (TNF-T), when synthesizing in strain-producer of Escherichia coli SG200-50 with plasmid pThy315, was a part of 'inclusion bodies' mostly in the form of a high-molecular complex with other proteins due to the S-S bonds formation. An approach of purification of TNF-T has been proposed, which is based on the destruction of the complex in the presence of sodium dodecylsulfate (DDS-Na) and dithiotreitol (DDT) followed by gel-filtration on Sephadex G-100 and renaturation by ultrafiltration on hollow fibers. The method allows the isolation of electrophoretically homogeneous TNF-T containing no DDS-Na and having high cytotoxic activity against cancer cells of mouse adenocarcinome L-929. The yield of TNF-T achieved 80% relative its content in biomass and 30% relative the total protein.
ISSN:0003-6838
1608-3024
DOI:10.1134/S0003683810020171